1. Field of the Invention
The present invention relates to a method of producing a minor ginsenoside using a ginsenoside glycosidase protein derived from a Microbacterium sp. microorganism, and a composition including the protein for conversion into a minor ginsenoside.
2. Description of the Related Art
Saponins, glycosides widely distributed in the plant kingdom, include diverse ring compounds formed by the non-sugar portion. Triterpene saponin, a saponin contained in ginseng or red ginseng as a major physiologically active ingredient, is named for distinguish with the other vegetable's saponin because of different chemical structure, called ginsenoside, which means ginseng glycoside.
Ginsenosides are classified into three groups based on their aglycone structure: Protopanaxadiol-type (PPD-type) ginsenosides, Protopanaxatriol-type (PPT-type) ginsenosides, and Oleanolic acid-type ginsenosides. These three groups are further classified based on the position and number of sugar moieties (aglycones) attached by a glycosidic bond at C-3, C-6, and C-20 positions of the rings in the chemical structure. PPD-type ginsenosides include Rb1, Rb2, Rb3, Rc, Rd, Gypenoside (Gyp) XVII, Compound O, Compound Mc1, F2, Compound Y, Compound Mc, Rg3, Rh2, and C-K. PPT-type ginsenosides include Re, Rg1, Rf, Rg2, Rh1 and F1.
In addition, major ginsenosides account for over 90% of total ginsenoside content in dry ginseng, but show a very low in-vivo absorption because of their large size of about 1,000 daltons. Therefore, in order to increase the efficacy of ginsenoside, it is required that major ginsenosides are converted into minor ginsenosides showing a relatively excellent absorption and efficacy. That is, deglycosylation of major ginsenosides for removing glucose, arabinose, rhamnose, xylose, etc. constituting sugars is required to show effective physiological activities in-vivo. The major ginsenosides include Rg1, Re, Rb1, Rb2, Rc, Rd or the like, and the minor ginsenosides (rare ginsenosides) present in trace amounts include F2, Rg3, Rh1, Rh2, gypenoside (Gyp) XVII, gypenoside LXXV, Compound K, C-K, Compound Mc, Compound Mc1 or the like.
Rare ginsenoside F1 is known to have efficacies of anti-aging and anti-oxidation, to protect HaCaT keratinocytes from UVB-induced cell apoptosis, and to have a skin whitening effect and an anti-cancer activity. Despite the usefulness of ginsenoside F1, its pharmacological activities have been recently reported, since it was found in 1976, because ginsenoside F1 is only present in the leaves of ginseng at relatively low concentrations, and thus it is difficult to obtain ginsenoside F1 in an amount sufficient for a biological activity test.
For the production of minor ginsenoside F1 present in ginseng in a small amount, a chemical decomposition, an enzymatic method, and a glycoside synthesis have been suggested, but these methods have limitations in mass-production, such as 1) many production steps required for the production process, 2) loss of desired compounds during processing, 3) use of inedible catalysts, or 4) low yield. In particular, with regard to the enzymatic method, there have many studies on use of coenzymes derived from various microorganisms and biotransformation of major ginsenosides of microorganisms. However, these methods are also not effective for mass-production, and have a problem of high production costs.
The production methods of F1 reported until now include biological production of several mg of F1, Rh1, Rg2 and Rg1 from 0.5 g of a PPT-type mixture by using glycosidase or production of F1 from Rg1 by using β-glucosidase isolated from fungus. Further, as the latest technology of producing a large amount of ginsenoside F1, it was reported that a large amount of F1 is successfully produced in grams by using β-glucosidase derived from a Terrabacter sp. microorganism (Korean Patent NO. 10-1340079).
Accordingly, the present inventors have made many efforts to develop a method of producing minor ginsenosides which are present in a trace amount in plants such as ginseng, etc. As a result, they found that more excellent bioconversion of major ginsenosides into minor ginsenosides is catalyzed by ginsenoside glycosidase MT619 derived from a Microbacterium sp. strain than β-glucosidase derived from a Terrabacter sp., thereby completing the present invention.